This research plan involves many aspects of central cholinergic neurons. The first section deals with localizing the muscarinic cholinergic receptor by light autoradiographic methods. This procedure will involve injecting rats with tritiated quinuclidinyl benzilate (QNB) which is a potent muscarinic cholinergic antagonist. Preliminary data indicate that the in vivo binding of QNB is at a pharmacologically relevant muscarinic cholinergic receptor. The autoradiographic localization of the QNB-binding sites will be performed utilizing established procedures which prevent diffusion artifacts. The second section of this study will examine various basic aspects of the recently discovered high affinity uptake mmchanism for choline which is localized to cholinergic nerve terminals. These studies involve: a critical reexamination of the kinetic constants for this uptake; an investigation of any significant exchange of the endogenous pools of synaptosomal choline with exogenous choline occurs during the in vitro uptake process; an examination of the structure-activity relationships of the choline uptake process which will provide some insight to the molecular and structural requirements of the choline uptake mechanism. The third section involves biochemical studies on cholinergic neuronal terminals after the neuronal impulse flow to these terminals has been altered. The neuronal impulse flow will be altered by axotomy which interrupts and stops impulse flow, and by electrical stimulation of the axons which will increase impulse flow. The biochemical studies will involve the metabolism of choline and acetylcholine and the subcellular distribution of choline and acetylcholine after these changes in neuronal impulse flow. The fourth section critically tests the relationship between acetylcolinesterase and cholinergic neurons within the hippocampus. There is evidence to suggest that in the hippocampus acetylcholinesterase is a specific marker for cholinergic neurons. This will be tested by examining if acetylcholinesterase is localized to neuronal terminals that appear to be noradrenergic or serotonergic. The noradrenergic and serotonergic neuronal terminals will be localized by autoradiographic methods. BIBLIOGRAPHIC REFERENCES: Simon, J. R. and Kuhar, M. J. (1976) High affinity choline uptake: Ionic and energy requirements. J. Neurochem. 25, in press